Project Summary. Regulated proteolysis controls the quality and quantity of proteins. In bacteria, energy dependent proteases eliminate aberrant proteins by recognizing distinctive marks arising from failed quality control, such as inappropriately exposed hydrophobic regions of proteins or specific tags attached upon prolonged translational arrest. These same machines also control levels and dynamics of fully active, well-folded proteins in order to properly manage molecular processes ranging from cell cycle progression to DNA damage. In this proposed work, we will focus on two major proteases systems that are found throughout bacteria and are crucial for virulence in human pathogens. First, we will determine how a newly discovered adaptor hierarchy controls the delivery of key proteins to the ClpXP protease during bacteria development and growth using structural, biochemical and genetic strategies. Second, we will uncover how the Lon protease, long known for its role in protein quality control, regulates stress responses and morphological changes building using systems-level approaches. By deciphering how bacteria govern the specificity and activity of these proteases at a mechanistic and cellular level, we will gain critical insight into a pathway that can be targeted by much needed new antibiotic strategies.